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Mutations in PLK4, encoding a master regulator of centriole biogenesis, cause microcephaly, growth failure and retinopathy

Abstract

Centrioles are essential for ciliogenesis. However, mutations in centriole biogenesis genes have been reported in primary microcephaly and Seckel syndrome, disorders without the hallmark clinical features of ciliopathies. Here we identify mutations in the genes encoding PLK4 kinase, a master regulator of centriole duplication, and its substrate TUBGCP6 in individuals with microcephalic primordial dwarfism and additional congenital anomalies, including retinopathy, thereby extending the human phenotypic spectrum associated with centriole dysfunction. Furthermore, we establish that different levels of impaired PLK4 activity result in growth and cilia phenotypes, providing a mechanism by which microcephaly disorders can occur with or without ciliopathic features.

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Figure 1: Individuals with mutations in PLK4 or TUBGCP6 exhibit extreme microcephaly and short stature.
Figure 2: Consequences of PLK4 mutations on the transcript and protein.
Figure 3: PLK4 mutations impair PLK4 activity in centriole biogenesis, resulting in reduced centriole number in patient-derived cells.
Figure 4: Depletion of plk4 causes dwarfism in zebrafish.
Figure 5: Impaired mitosis leads to growth retardation in plk4-morphant zebrafish.
Figure 6: plk4-morphant zebrafish display retinal defects due to reduced cilia number.
Figure 7: The growth failure and ciliopathy phenotypes are separable in a dose-dependent manner.

References

  1. 1

    Nigg, E.A. & Raff, J.W. Centrioles, centrosomes, and cilia in health and disease. Cell 139, 663–678 (2009).

    CAS  Article  Google Scholar 

  2. 2

    Bettencourt-Dias, M., Hildebrandt, F., Pellman, D., Woods, G. & Godinho, S.A. Centrosomes and cilia in human disease. Trends Genet. 27, 307–315 (2011).

    CAS  Article  Google Scholar 

  3. 3

    Sir, J.H. et al. Loss of centrioles causes chromosomal instability in vertebrate somatic cells. J. Cell Biol. 203, 747–756 (2013).

    CAS  Article  Google Scholar 

  4. 4

    Ganem, N.J., Godinho, S.A. & Pellman, D. A mechanism linking extra centrosomes to chromosomal instability. Nature 460, 278–282 (2009).

    CAS  Article  Google Scholar 

  5. 5

    Gönczy, P. Towards a molecular architecture of centriole assembly. Nat. Rev. Mol. Cell Biol. 13, 425–435 (2012).

    Article  Google Scholar 

  6. 6

    Guernsey, D.L. et al. Mutations in centrosomal protein CEP152 in primary microcephaly families linked to MCPH4. Am. J. Hum. Genet. 87, 40–51 (2010).

    CAS  Article  Google Scholar 

  7. 7

    Kalay, E. et al. CEP152 is a genome maintenance protein disrupted in Seckel syndrome. Nat. Genet. 43, 23–26 (2011).

    CAS  Article  Google Scholar 

  8. 8

    Bond, J. et al. A centrosomal mechanism involving CDK5RAP2 and CENPJ controls brain size. Nat. Genet. 37, 353–355 (2005).

    CAS  Article  Google Scholar 

  9. 9

    Kumar, A., Girimaji, S.C., Duvvari, M.R. & Blanton, S.H. Mutations in STIL, encoding a pericentriolar and centrosomal protein, cause primary microcephaly. Am. J. Hum. Genet. 84, 286–290 (2009).

    CAS  Article  Google Scholar 

  10. 10

    Sir, J.H. et al. A primary microcephaly protein complex forms a ring around parental centrioles. Nat. Genet. 43, 1147–1153 (2011).

    CAS  Article  Google Scholar 

  11. 11

    Hussain, M.S. et al. A truncating mutation of CEP135 causes primary microcephaly and disturbed centrosomal function. Am. J. Hum. Genet. 90, 871–878 (2012).

    CAS  Article  Google Scholar 

  12. 12

    Bettencourt-Dias, M. et al. SAK/PLK4 is required for centriole duplication and flagella development. Curr. Biol. 15, 2199–2207 (2005).

    CAS  Article  Google Scholar 

  13. 13

    Habedanck, R., Stierhof, Y.D., Wilkinson, C.J. & Nigg, E.A. The Polo kinase Plk4 functions in centriole duplication. Nat. Cell Biol. 7, 1140–1146 (2005).

    CAS  Article  Google Scholar 

  14. 14

    Verloes, A., Drunat, S., Gressens, P. & Passemard, S. in GeneReviews (eds. Pagon, R.A. et al.) 1–2 (University of Washington, Seattle, 1993).

  15. 15

    Bahtz, R. et al. GCP6 is a substrate of Plk4 and required for centriole duplication. J. Cell Sci. 125, 486–496 (2012).

    CAS  Article  Google Scholar 

  16. 16

    Puffenberger, E.G. et al. Genetic mapping and exome sequencing identify variants associated with five novel diseases. PLoS ONE 7, e28936 (2012).

    CAS  Article  Google Scholar 

  17. 17

    Hudson, J.W. et al. Late mitotic failure in mice lacking Sak, a polo-like kinase. Curr. Biol. 11, 441–446 (2001).

    CAS  Article  Google Scholar 

  18. 18

    Cizmecioglu, O. et al. Cep152 acts as a scaffold for recruitment of Plk4 and CPAP to the centrosome. J. Cell Biol. 191, 731–739 (2010).

    CAS  Article  Google Scholar 

  19. 19

    Kleylein-Sohn, J. et al. Plk4-induced centriole biogenesis in human cells. Dev. Cell 13, 190–202 (2007).

    CAS  Article  Google Scholar 

  20. 20

    Hornick, J.E. et al. Amphiastral mitotic spindle assembly in vertebrate cells lacking centrosomes. Curr. Biol. 21, 598–605 (2011).

    CAS  Article  Google Scholar 

  21. 21

    Bicknell, L.S. et al. Mutations in ORC1, encoding the largest subunit of the origin recognition complex, cause microcephalic primordial dwarfism resembling Meier-Gorlin syndrome. Nat. Genet. 43, 350–355 (2011).

    CAS  Article  Google Scholar 

  22. 22

    Pfaff, K.L. et al. The zebrafish cassiopeia mutant reveals that SIL is required for mitotic spindle organization. Mol. Cell. Biol. 27, 5887–5897 (2007).

    CAS  Article  Google Scholar 

  23. 23

    Coelho, P.A. et al. Spindle formation in the mouse embryo requires Plk4 in the absence of centrioles. Dev. Cell 27, 586–597 (2013).

    CAS  Article  Google Scholar 

  24. 24

    Izraeli, S. et al. The SIL gene is required for mouse embryonic axial development and left-right specification. Nature 399, 691–694 (1999).

    CAS  Article  Google Scholar 

  25. 25

    Bazzi, H. & Anderson, K.V. Acentriolar mitosis activates a p53-dependent apoptosis pathway in the mouse embryo. Proc. Natl. Acad. Sci. USA 111, E1491–E1500 (2014).

    CAS  Article  Google Scholar 

  26. 26

    McIntyre, R.E. et al. Disruption of mouse Cenpj, a regulator of centriole biogenesis, phenocopies Seckel syndrome. PLoS Genet. 8, e1003022 (2012).

    CAS  Article  Google Scholar 

  27. 27

    Larison, K.D. & Bremiller, R. Early onset of phenotype and cell patterning in the embryonic zebrafish retina. Development 109, 567–576 (1990).

    CAS  PubMed  Google Scholar 

  28. 28

    Wheway, G., Parry, D.A. & Johnson, C.A. The role of primary cilia in the development and disease of the retina. Organogenesis 10, 69–85 (2014).

    Article  Google Scholar 

  29. 29

    Blachon, S. et al. Drosophila asterless and vertebrate Cep152 are orthologs essential for centriole duplication. Genetics 180, 2081–2094 (2008).

    CAS  Article  Google Scholar 

  30. 30

    Slevin, L.K. et al. The structure of the plk4 cryptic polo box reveals two tandem polo boxes required for centriole duplication. Structure 20, 1905–1917 (2012).

    CAS  Article  Google Scholar 

  31. 31

    Novorol, C. et al. Microcephaly models in the developing zebrafish retinal neuroepithelium point to an underlying defect in metaphase progression. Open Biol. 3, 130065 (2013).

    Article  Google Scholar 

  32. 32

    Sillibourne, J.E. & Bornens, M. Polo-like kinase 4: the odd one out of the family. Cell Div. 5, 25 (2010).

    Article  Google Scholar 

  33. 33

    Ko, M.A. et al. Plk4 haploinsufficiency causes mitotic infidelity and carcinogenesis. Nat. Genet. 37, 883–888 (2005).

    CAS  Article  Google Scholar 

  34. 34

    Holland, A.J. et al. Polo-like kinase 4 controls centriole duplication but does not directly regulate cytokinesis. Mol. Biol. Cell 23, 1838–1845 (2012).

    CAS  Article  Google Scholar 

  35. 35

    Holland, A.J., Lan, W., Niessen, S., Hoover, H. & Cleveland, D.W. Polo-like kinase 4 kinase activity limits centrosome overduplication by autoregulating its own stability. J. Cell Biol. 188, 191–198 (2010).

    CAS  Article  Google Scholar 

  36. 36

    Lancaster, M.A. & Knoblich, J.A. Spindle orientation in mammalian cerebral cortical development. Curr. Opin. Neurobiol. 22, 737–746 (2012).

    CAS  Article  Google Scholar 

  37. 37

    Chen, J.F. et al. Microcephaly disease gene Wdr62 regulates mitotic progression of embryonic neural stem cells and brain size. Nat. Commun. 5, 3885 (2014).

    CAS  Article  Google Scholar 

  38. 38

    Thompson, S.L. & Compton, D.A. Examining the link between chromosomal instability and aneuploidy in human cells. J. Cell Biol. 180, 665–672 (2008).

    CAS  Article  Google Scholar 

  39. 39

    Marthiens, V. et al. Centrosome amplification causes microcephaly. Nat. Cell Biol. 15, 731–740 (2013).

    CAS  Article  Google Scholar 

  40. 40

    Hanks, S. et al. Constitutional aneuploidy and cancer predisposition caused by biallelic mutations in BUB1B. Nat. Genet. 36, 1159–1161 (2004).

    CAS  Article  Google Scholar 

  41. 41

    Sorokin, S.P. Reconstructions of centriole formation and ciliogenesis in mammalian lungs. J. Cell Sci. 3, 207–230 (1968).

    CAS  Google Scholar 

  42. 42

    Shinohara, K. et al. Two rotating cilia in the node cavity are sufficient to break left-right symmetry in the mouse embryo. Nat. Commun. 3, 622 (2012).

    Article  Google Scholar 

  43. 43

    Li, H. & Durbin, R. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics 25, 1754–1760 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  44. 44

    McLaren, W. et al. Deriving the consequences of genomic variants with the Ensembl API and SNP Effect Predictor. Bioinformatics 26, 2069–2070 (2010).

    CAS  Article  Google Scholar 

  45. 45

    McKenna, A. et al. The Genome Analysis Toolkit: a MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res. 20, 1297–1303 (2010).

    CAS  Article  Google Scholar 

  46. 46

    Yeo, G. & Burge, C.B. Maximum entropy modeling of short sequence motifs with applications to RNA splicing signals. J. Comput. Biol. 11, 377–394 (2004).

    CAS  Article  Google Scholar 

  47. 47

    Gudbjartsson, D.F., Jonasson, K., Frigge, M.L. & Kong, A. Allegro, a new computer program for multipoint linkage analysis. Nat. Genet. 25, 12–13 (2000).

    CAS  Article  Google Scholar 

  48. 48

    Hussain, M.S. et al. CDK6 associates with the centrosome during mitosis and is mutated in a large Pakistani family with primary microcephaly. Hum. Mol. Genet. 22, 5199–5214 (2013).

    CAS  Article  Google Scholar 

  49. 49

    Livak, K.J. & Schmittgen, T.D. Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 25, 402–408 (2001).

    CAS  Article  Google Scholar 

  50. 50

    Wolff, A. et al. Distribution of glutamylated α and β-tubulin in mouse tissues using a specific monoclonal antibody, GT335. Eur. J. Cell Biol. 59, 425–432 (1992).

    CAS  PubMed  Google Scholar 

  51. 51

    Kimmel, C.B., Ballard, W.W., Kimmel, S.R., Ullmann, B. & Schilling, T.F. Stages of embryonic development of the zebrafish. Dev. Dyn. 203, 253–310 (1995).

    CAS  Google Scholar 

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Acknowledgements

We thank the families and clinicians for their involvement and participation; P. Mills, T. Hurd, M. Bettencourt-Dias and M. Reijns for commenting on the manuscript; N. Hastie, D. Fitzpatrick and J. Livingston for helpful discussions; C. Janke (Institut Curie) for his kind gift of the GT335 antibody; E. Freyer for assistance with FACS analysis; P. Gautier for bioinformatics; P. Carroll and A. Vickers for technical assistance; the IGMM core sequencing service; the IGMM imaging facility for assistance with microscopy; E. Patton and the IGMM fish facility for advice and zebrafish technical assistance; E. Liston and the DNA Resource Centre at SickKids for sample processing; A. Pearce and E. Maher (Cytogenetics Laboratory, South East Scotland Genetics Service) for technical advice; G. Hahn (University Hospital Carl Gustav Carus) for her second opinion on the MRI data; and N. Dalibor and E. Kirst (CCG) for their expert technical assistance. This work was supported by funding from the MRC, the Lister Institute for Preventative Medicine and the European Research Council (ERC, 281847) (A.P.J.), Medical Research Scotland (L.S.B.), the National Institute for Health Research Moorfields Eye Hospital Biomedical Research Centre (A.T.M.), Köln Fortune (M.S.H.) and CMMC (P.N. and A.A.N.).

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P.N., H.T., J.A., M.S.H., A.B., K.M., M.E. Hurles, J.E.M. and L.S.B. performed exome sequencing and analysis. L.S.B., C.-A.M., J.E.M., M.R.T., I.A., M.S.H. and G.N. performed sequencing, genotyping, linkage analysis and other molecular genetics experiments. C.-A.M., A.L., C.K., M.E. Harley, I.A., M.S.H., R.M., A.A.N. and I.H. designed and performed the cell biology experiments. A. Klingseisen designed and performed the zebrafish experiments with help from A.L., C.-A.M., J.D. and P.H. W.H. performed structural analysis. D.H., F.K., Z.A., S.T., V.C.-D., H.D., L.D., A. Kariminejad, R.M.-L., A.T.M., A.S., C.S., R.W. and S.M.B. ascertained subjects, obtained samples and/or assisted with phenotypic analysis and clinical studies. C.-A.M. and A.P.J. wrote the manuscript with help from P.N., A. Klingseisen and L.S.B. The study was planned and supervised by P.N. and A.P.J.

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Correspondence to Peter Nürnberg or Andrew P Jackson.

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P.N. is a founder, CEO and shareholder of ATLAS Biolabs. ATLAS Biolabs is a service provider for genomic analyses.

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Martin, CA., Ahmad, I., Klingseisen, A. et al. Mutations in PLK4, encoding a master regulator of centriole biogenesis, cause microcephaly, growth failure and retinopathy. Nat Genet 46, 1283–1292 (2014). https://doi.org/10.1038/ng.3122

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